title: Molecular cloning and characterization of cystatin, a cysteine protease inhibitor, from bufo melanostictus.
abstract: Cystatins are efficient inhibitors of papain-like cysteine proteinases, and they serve various important physiological functions. In this study, a novel cystatin, Cystatin-X, was cloned from a cDNA library of the skin of Bufo melanostictus. The single nonglycosylated polypeptide chain of Cystatin-X consisted of 102 amino acid residues, including seven cysteines. Evolutionary analysis indicated that Cystatin-X can be grouped with family 1 cystatins. It contains cystatin-conserved motifs known to interact with the active site of cysteine proteinases. Recombinant Cystatin-X expressed and purified from Escherichia coli exhibited obvious inhibitory activity against cathepsin B. rCystatin-X at a concentration of 8 µM inhibited nearly 80% of cathepsin B activity within 15 s, and about 90% of cathepsin B activity within 15 min. The Cystatin-X identified in this study can play an important role in host immunity and in the medical effect of B. melanostictus.
authors.affiliation: Engineering Research Center for Molecular Diagnosis, Faculty of Life Science and Technology, Kunming University of Science and Technology.
authors.forename: Wa
authors.initials: W
authors.lastname: Liu
authors.forename: Senlin
authors.initials: S
authors.lastname: Ji
authors.forename: A-Mei
authors.initials: AM
authors.lastname: Zhang
authors.forename: Qinqin
authors.initials: Q
authors.lastname: Han
authors.forename: Yue
authors.initials: Y
authors.lastname: Feng
authors.forename: Yuzhu
authors.initials: Y
authors.lastname: Song
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002025:Bufonidae / Q000235:genetics*; D002401:Cathepsin B / Q000378:metabolism; D003001:Cloning, Molecular; D015891:Cystatins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D017421:Sequence Analysis
chemical_list: D015891:Cystatins; D015853:Cysteine Proteinase Inhibitors; D002401:Cathepsin B